慢病毒介导的微小RNA-191对小鼠视网膜新生血管的抑制作用

miR-191 inhibits oxygen-induced retinal neovascularization in mice

目的观察慢病毒介导(LV)的微小RNA (miR)-191对氧诱导视网膜病变(OIR)模型小鼠视网膜新生血管(RNV)的抑制作用。方法7日龄C57BL/6J小鼠80只,随机分为正常组、非干预组、LV miR-191 (LV-191)组、生理盐水(NS)组、LV-绿色荧光蛋白(GFP)组,每组均为16只。参照文献方法建立OIR小鼠模型。12日龄时,LV-191组、NS组、LV-GFP组,小鼠右眼玻璃体腔分别注射1 μl LV-191、NS、LV-GFP;正常组、非干预组小鼠不做处理。视网膜铺片观察视网膜无灌注区相对面积,病理切片计数突破内界膜的视网膜血管内皮细胞核数;实时定量PCR (RT-PCR)检测小鼠视网膜中LV-191、p21 mRNA相对表达量。结果正常组、非干预组、LV-191组、NS组、LV-GFP组小鼠视网膜无灌注区面积比较,差异有统计学意义(F=127.20,P<0.001);突破内界膜的血管内皮细胞核计数比较,差异有统计学意义(F=31.71 ,P<0.05)。RT-PCR检测结果显示,正常组、非干预组、LV-191组、NS组、LV-GFP组小鼠视网膜中miR-191、p21 mRNA相对表达量比较,差异有统计学意义(F=10.95、15.60,P<0.05、<0.05)。与正常组、非干预组、NS组、LV-GFP组小鼠比较,LV-191组视网膜中miR-191、p21 mRNA相对表达量均明显上调,差异有统计学意义(P<0.05)。结论慢病毒介导的miR-191可通过上调p21表达抑制RNV的生成。

Objective To observe the inhibitory effect of lentiviral vector miR-191 (LV-191) on retinal neovascularization (RNV) in mice model of oxygen-induced retinopathy (OIR). Methods Eighty healthy 7-day-old C57BL/6J mice were randomly divided into 5 groups including normal group, non-intervention group, normal saline (NS) group, LV-191 group and LV-green fluorescent protein (GFP) group, 16 mice in each group. The OIR model was established in the non-intervention group, NS group, LV-191 group and LV-GFP group. NS group, LV-191 group and LV-GFP group were given an intravitreal injection of 1 μl of NS, LV-191 and LV-GFP at the age of 12 days. No injection was performed in the non-intervention group. In normal group,newborn mouse were maintained in room air form P0 to P17, and no treatment was performed. Mice in all five groups were euthanized at P17. Retinal neovasculation (RNV) was evaluated by counting the number of pre-retinal neovascular cells and analysis of non-perfusion area area by immunofluorescent staining of the mouse retina. Real-time quantitative PCR (RT-PCR) to detect miR -191 and P21 expression of retinal tissue. Results In the LV-191 group, the non-perfusion area were both significantly smaller than those in non-intervention group, NS group and LV-GFP group (F=127.20, P<0.001). The number of pre-retinal neovascular cell nuclei in retinas from LV-191 group were obviously lower than those in the retinas from non-intervention group, NS group and LV-GFP group (F=31.71, P<0.05). RT-PCR showed that the LV-191 and P21 level of LV-191 group increased significantly than other groups (F=10.95, 15.60;P<0.05). Conclusion Intravitreal injection of LV- 191 inhibits RNV in mice model of OIR possibly through up-regulating p21.